OBJECTIVE: To analyze the mutations in transcription regulatory sequences (TRSs) of coronaviruss (CoV) to provide the basis for exploring the patterns of SARS-CoV-2 transmission and outbreak. METHODS: A combined evolutionary and molecular functional analysis of all sets of publicly available genomic data of viruses was performed. RESULTS: A leader transcription regulatory sequence (TRS-L) usually comprises the first 60-70 nts of the 5' UTR in a CoV genome, and the body transcription regulatory sequences (TRS-Bs) are located immediately upstream of the genes other than ORF1a and 1b. In each CoV genome, the TRS-L and TRS-Bs share a specific consensus sequence, namely the TRS motif. Any changes of nucleotide residues in the TRS motifs are defined as TRS motif mutations. Mutations in the TRS-L or multiple TRS-Bs result in superattenuated variants. The spread of super-attenuated variants may cause an increase in asymptomatic or mild infections, prolonged incubation periods and a decreased detection rate of the viruses, thus posing new challenges to SARS-CoV-2 prevention and control. The super-attenuated variants also increase their possibility of long-term coexistence with humans. The Delta variant is significantly different from all the previous variants and may lead to a large-scale transmission. The Delta variant (B.1.617.2) with TRS motif mutation has already appeared and shown signs of spreading in Singapore, which, and even the Southeast Asia, may become the new epicenter of the next wave of SARS-CoV-2 outbreak. CONCLUSION TRS motif mutation will occur in all variants of SARS-CoV-2 and may result in super-attenuated variants. Only super-attenuated variants with TRS motif mutations will eventually lose the abilities of cross-species transmission and causing outbreaks.
COVID-19/virology* ; Genome, Viral ; Humans ; Mutation ; SARS-CoV-2/genetics*

The aims of this study were to determine the prognostic value of primary tumor surgery and identify optimal candidates for such surgery among patients with seminoma and distant metastasis at diagnosis. We identified 521 patients with seminoma and distant metastasis at diagnosis between 2004 and 2014 from the Surveillance, Epidemiology, and End Results database. Among these patients, 434 had undergone surgery, whereas 87 had not. The prognostic value of primary tumor surgery was assessed by Kaplan-Meier methods, log-rank analyses, and multivariate Cox's proportional hazards model. Survival curves and forest plots were also plotted. Survival analysis indicated that patients who underwent surgery had a better 5-year overall survival and cancer-specific survival than those who did not. Multivariate analyses demonstrated that primary tumor surgery is an independent prognostic factor for overall survival and cancer-specific survival, along with age at diagnosis, M stage, and marital status. In addition, primary tumor surgery still had considerable prognostic value in the subgroup of patients with lymph node metastasis. Further, forest plots demonstrated that patients with M1a stage, N1 or N2-3 stage, and a younger age at diagnosis (<60 years) may benefit from primary tumor surgery. In conclusion, our findings indicate that primary tumor surgery is correlated with improved survival in patients with seminoma and distant metastasis. Furthermore, primary tumor surgery is an independent prognostic indicator for patients with seminoma and distant metastasis.

OBJECTIVE: To investigate the current status of antibiotic use for very and extremely low birth weight (VLBW/ELBW) infants in neonatal intensive care units (NICUs) of Hunan Province. METHODS: The use of antibiotics was investigated in multiple level 3 NICUs of Hunan Province for VLBW and ELBW infants born between January, 2017 and December, 2017. RESULTS: The clinical data of 1 442 VLBW/ELBW infants were collected from 24 NICUs in 2017. The median antibiotic use duration was 17 days (range: 0-86 days), accounting for 53.0% of the total length of hospital stay. The highest duration of antibiotic use was up to 91.4% of the total length of hospital stay, with the lowest at 14.6%. In 16 out of 24 NICUs, the antibiotic use duration was accounted for more than 50.0% of the hospitalization days. There were 113 cases with positive bacterial culture grown in blood or cerebrospinal fluid, making the positive rate of overall bacterial culture as 7.84%. The positive rate of bacterial culture in different NICUs was significantly different from 0% to 14.9%. The common isolated bacterial pathogens Klebsiella pneumoniae was 29 cases (25.7%); Escherichia coli 12 cases (10.6%); Staphylococcus aureus 3 cases (2.7%). The most commonly used antibiotics were third-generation of cephalosporins, accounting for 41.00% of the total antibiotics, followed by penicillins, accounting for 32.10%, and followed by carbapenems, accounting for 13.15%. The proportion of antibiotic use time was negatively correlated with birth weight Z-score and the change in weight Z-score between birth and hospital discharge (r=-0.095, -0.151 respectively, P<0.01), positively correlated with death/withdrawal of care (r=0.196, P<0.01). CONCLUSIONS Antibiotics used for VLBW/ELBW infants in NICUs of Hunan Province are obviously prolonged in many NICUs. The proportion of routine use of third-generation of cephalosporins and carbapenems antibiotics is high among the NICUs.
Anti-Bacterial Agents ; Birth Weight ; Humans ; Infant ; Infant, Extremely Low Birth Weight ; Infant, Newborn ; Intensive Care Units, Neonatal ; Surveys and Questionnaires

AIM:To analyze the imageological changes of acute and chronic central serous chorioretinopathy (CSC) by 2 types of optical coherence tomography (OCT). METHODS:A retrospective analysis was performed, inclu-ding data of 60 eyes from 56 patients with CSC diagnosed by conventional eye examination, fundus fluorescein angiography (FFA) and indocyanine green angiography (ICGA), which were divided into acute group (28 eyes of 28 patients) and chronic group (32 eyes of 28 patients) according to imageological examinations and duration (6 months). Optical coher-ence tomography angiography (OCTA) and spectral domain optical coherence tomography ( SD-OCT) were performed to study the vessel density of the chorioretinal leyers and the integrity of the outer retinal structure. RESULTS:In the pa-tients with chronic CSC, OCTA in 4 eyes ( 12.50% ) revealed the presence of a distinct choroidal neovascularization (CNV), while no evidence of CNV in ICGA was observed. However, no sign of CNV in acute CSC group both on OCTA and ICGA was found. The occurrence of 'dark areas' in chronic CSC was much higher than that in acute CSC ( P <0.01). In addition, the integrity of the outer retinal structure (defined as tissue between external limiting membrane and retinal pigment epithelium) in acute group was significantly better than that in chronic group ( P <0.01). CONCLU-SION:Our study demonstrates the existing secondary CNV that is not demonstrated by ICGA in the chronic CSC patients, and the different characteristics of retinochoroid structures between acute and chronic CSC in OCTA and SD-OCT are ob- served. Chronic CSC has more severe structural changes.

The aim of this paper was to explore the effects and possible mechanisms in vitro of tea polyphenols (TP) delaying human glomerular mesangial cells (HGMCs) senescence induced by high glucose (HG). HGMCs were cultured in vitro and divided into the normal group (N, 5.5 mmol·L⁻¹ glucose), the mannitol group(MNT, 5.5 mmol·L⁻¹ glucose plus 24.5 mmol·L⁻¹ mannitol), the high dose of D-glucose group (HG, 30 mmol·L⁻¹ glucose), the low dose of TP group (L-TP, 30 mmol·L⁻¹ glucose plus 5 mg·L⁻¹ TP) and the high dose of TP group (H-TP, 30 mmol·L⁻¹ glucose plus 20 mg·L⁻¹ TP), which were cultured in 5% CO₂ at 37 °C, respectively. Firstly, the effects of TP on the cell morphology of HGMCs were observed after 72 h-intervention. Secondly, the cell cycle, the positive rate of senescence-associated-β-galactosidase (SA-β-gal) staining and the telomere length were detected, respectively. Finally, the protein expressions of p53, p21 and Rb in the p53-p21-Rb signaling pathway were investigated, respectively. And the expressions of p-STAT3 and miR-126 were examined severally. The results indicated that HG not only arrested the cell cycle in G₁ phase but also increased the positive rate of SA-β-gal staining, and shortened the telomere length. HG led to the protein over-expressions of p53, p21 and Rb and HGMCs senescence by activating the p53-p21-Rb signaling pathway. In addition, L-TP delayed HGMCs senescence by improving the cell cycle G₁ arrest, reducing SA-β-gal staining positive rate and lengthening the telomere length. L-TP reduced the protein over-expressions of p53, P21 and Rb induced by HG and inhibited the telomere-p53-p21-Rb signaling pathway. Moreover, the expression of p-STAT3 was increased and the expression of miR-126 was decreased in HGMCs induced by HG. L-TP reduced the expression of p-STAT3 and increased the expression of miR-126 in HGMCs. In conclusion, HG could induce HGMCs senescence by activating the telomere-p53-p21-Rb signaling pathway in vitro. L-TP could delay HGMCs senescence through regulating STAT3/miR-126 expressions and inhibiting the telomere-p53-p21-Rb signaling pathway activation. These findings could provide the effective interventions in clinic for preventing and treating renal cell senescence in diabetic kidney disease.
Cells, Cultured ; Cellular Senescence ; Cyclin-Dependent Kinase Inhibitor p21 ; Glucose ; Humans ; Mesangial Cells ; MicroRNAs ; Polyphenols ; STAT3 Transcription Factor ; Tea ; Telomere ; Tumor Suppressor Protein p53

Objective:To investigate the clinical usefulness of echocardiography in the diagnosis of infracardiac total anomalous pulmonary venous connection (ITAPVC) in neonates and infants.Methods:Retrospective analysis on 8 patients with ITAPVC was performed using echocardiography between April 2006 and December 2016.There were 4 boys and 4 girls with a mean age of 79.8 days (ranging from 15 to 195 days).A combined scanning via parasternal,subcostal and apical acoustic windows had been employed to diagnose ITAPVC and to trace the course and site of the anomalous pulmonary venous drainage,and to confirm the direction of the inter-atrial shunt and enlargement of right atrium and right ventricle.Results:Of the 8 patients who received echocardiography,ITAPVC was diagnosed in 7 patients.Misdiagnosis by echocardiography was encountered in one patient.The diagnosis by echocardiography was compatible with the operative findings in 5 patients receiving surgery and with the results of multislice computed tomography in 6 patients.The diagnostic accuracy rate of ITAPVC was 87.5％.The indirect signs obtained from echocardiogram was coexistence of a small malformed,triangle-shaped left atrium and right to left shunting at atrial level with dilatation and tortuousness of portal vein or hepatic vein and abundant blood flow in liver.The direct signs was total pulmonary veins unconnected with left atrium,whose confluence joining into vertical vein drained right-inferiorly to portal vein or hepatic vein through diaphragm.Three parallel vessels including vertical vein,abdominal aorta and inferior vena cava arranged anteriorly,left-posteriorly and right-posteriorly with the opposite flow directions of inferior vena cava and the other two were found on sub-costal view.Sites of the drainage to the infra-diaphragm veins located portal vein in 8 patients.Stenosis of site of vertical vein connecting to portal vein or hepatic veins occurred in 3 patients.Conclusion:Echocardiography has significant value in the diagnosis of pediatric ITAPVC and is capable of providing important structural and hemodynamic information for preoperative assessment of surgery.With multiple windows and multiple sections,ITAPVC could be diagnosed accurately by echocardiography.However,it is necessary to differentiate ITAPVC with intrahepatic portosystemic venous shunts or hepatic arteriovenous fistula.

Aconiti Lateralis Radix Praeparata (Fuzi) is a commonly used traditional Chinese medicine in clinic for its potency in restoring yang and rescuing from collapse. Aconiti alkaloids, mainly including monoester-diterpenoidaconitines (MDAs) and diester-diterpenoidaconitines (DDAs), are considered to act as both bioactive and toxic constituents. In the present study, a feasible, economical, and accurate HPLC method for simultaneous determination of six alkaloid markers using the Single Standard for Determination of Multi-Components (SSDMC) method was developed and fully validated. Benzoylmesaconine was used as the unique reference standard. This method was proven as accurate (recovery varying between 97.5%-101.8%, RSD < 3%), precise (RSD 0.63%-2.05%), and linear (R > 0.999 9) over the concentration ranges, and subsequently applied to quantitative evaluation of 62 batches of samples, among which 45 batches were from good manufacturing practice (GMP) facilities and 17 batches from the drug market. The contents were then analyzed by principal component analysis (PCA) and homogeneity test. The present study provided valuable information for improving the quality standard of Aconiti Lateralis Radix Praeparata. The developed method also has the potential in analysis of other Aconitum species, such as Aconitum carmichaelii (prepared parent root) and Aconitum kusnezoffii (prepared root).
Aconitine ; analogs & derivatives ; chemistry ; Aconitum ; chemistry ; Alkaloids ; analysis ; Biomarkers ; analysis ; Chromatography, High Pressure Liquid ; economics ; methods ; Diterpenes ; chemistry ; Drugs, Chinese Herbal ; chemistry ; Feasibility Studies ; Molecular Structure

Bletilla striata has been used as traditional Chinese medicine for several centuries. In recent years, the quality and quantity of wild B. striata plants have declined sharply due to habitat deterioration and human over-exploitation. Therefore, it is of great urgency to evaluate and protect B. striata wild plant resource. In this study, sequence-related amplified polymorphism (SRAP) markers were applied to assess the level and pattern of genetic diversity in twelve populations of B. striata. The results showed a high level of genetic diversity (PPB = 90.48%, H = 0.349 4, I = 0.509 6) and moderate genetic differentiation among populations (G(st) = 0.260 9). Based on the unweighted pair-group method with arithmetic average (UPGMA), twelve populations gathered in three clusters. The cluster 1 included four populations. There are Nanjing, Zhenjiang, Xuancheng and Hangzhou. The seven populations which come from Hubei Province, Hunan Province, Jiangxi Province and Guizhou Province belonged to the cluster 2. The cluster 3 only contained Wenshan population. Moreover, Mantel test revealed significant positive correlation between genetic distances and geographic distances (r = 0.632 9; P < 0.000 1). According to the results, we proposed a series of conservation consideration for B. striata.
China ; Genetic Markers ; Genetic Variation ; Genetics, Population ; Orchidaceae ; genetics ; Phylogeny ; Plants, Medicinal ; genetics

Bletilla striata has been used as traditional Chinese medicine for several centuries. In recent years, the quality and quantity of wild B. striata plants have declined sharply due to habitat deterioration and human over-exploitation. Therefore, it is of great urgency to evaluate and protect B. striata wild plant resource. In this study, sequence-related amplified polymorphism (SRAP) markers were applied to assess the level and pattern of genetic diversity in twelve populations of B. striata. The results showed a high level of genetic diversity (PPB = 90.48%, H = 0.349 4, I = 0.509 6) and moderate genetic differentiation among populations (G_st = 0.260 9). Based on the unweighted pair-group method with arithmetic average (UPGMA), twelve populations gathered in three clusters. The cluster 1 included four populations. There are Nanjing, Zhenjiang, Xuancheng and Hangzhou. The seven populations which come from Hubei Province, Hunan Province, Jiangxi Province and Guizhou Province belonged to the cluster 2. The cluster 3 only contained Wenshan population. Moreover, Mantel test revealed significant positive correlation between genetic distances and geographic distances (r = 0.632 9; P < 0.000 1). According to the results, we proposed a series of conservation consideration for B. striata.

AIM:Toinvestigatetheeffectsofcalcitoningene-relatedpeptide(CGRP)onmyocardinexpres-sion and phenotypic switch in vascular smooth muscle cells ( VSMCs) .METHODS:VSMCs were obtained by aortic tissue adherent culture and treated with angiotensin Ⅱ( AngⅡ) , AngⅡ+CGRP or AngⅡ+CGRP +CGRP8-37 .The protein expression of myocardin and the phenotypic proteins of the VSMCs was detected by Western blot .RESULTS:The expres-sion of myocardin in cultured VSMCs showed downregulation along with time expansion .The protein level of myocardin was higher at 48 h and 72 h than that at baseline in the cultured VSMCs (P<0.05).However, the myocardin was lower at 48 h and 72 h than that at baseline after treatment with CGRP in cultured VSMCs (P<0.05).Furthermore, at 48 h in cul-tured VSMCs, the myocardin decreased along with α-smooth muscle actin (α-SMA) (P<0.05), and osteopontin (OPN) increased (P<0.05) in AngⅡ group compared with control group .After treatment with CGRP, the levels of myocardin andα-SMA become higher ( P <0.05 ) but OPN was lower ( P <0.05 ) in CGRP group than those in AngⅡ group. CGRP8-37 abrogated CGRP-induced increase in myocardin and α-SMA and decrease in OPN in CGRP 8-37 group compared with CGRP group .CONCLUSION: CGRP may regulate the phenotypic switch of the VSMCs and maintain the cells in contractile phenotype through the upregulation of myocardin protein , which may be accomplished by the combination of CGRP and its receptor .